Dengue virus is a human pathogenic flavivirus that has re-emerged as a serious public health threat. There is no treatment or vaccine, and public health control measures have failed. There is little understanding of the pathogenesis of dengue virus infection (or that of any other flavivirus). The binding of viral ectodomain molecules to specific receptors on target cells is a critical factor in pathogenicity as it is an important determinant of cell and tissue tropism; yet, there is no information about the molecular basis of the binding of dengue virus to target cells. We have characterized the interaction of dengue virus envelope protein with target cells, and partially identified a target cell receptor. We propose to fully identify the proteoglycan that is a cellular receptor for dengue virus, assess for the presence of co-receptor mechanisms, identify the envelope protein domains responsible for cell-binding, and develop inhibitors of infection based on blockade of viral target cell binding. Aim 1 will identify the proteoglycan (PG) that is the dengue receptor for Vero and other cells. Results indicate that the glycosaminoglycan (GAG) component of this PG embodies the receptor specificity, and is an unusually highly sulfated form of heparan sulfate. Aim 2 will identify the dengue envelope protein motifs responsible for target cell binding. Results indicate that the envelope protein accounts for viral binding, and has GAG-binding domains in two regions at the carboxy-terminus. Recombinant envelope proteins with mutations in these regions will be expressed, and their binding properties compared with wild-type envelope protein. Infectious virus incorporating mutations of interest will be generated, and assessed for changes in infectivity. Aim 3 will identify pharmaceuticals that prevent infection by blocking virus - target cell binding. Results indicate that the polysulfonate Suramin inhibits both envelope protein binding, and infectivity. Using the structure of Suramin as a guide, Suramin analogues and other sulfated/sialylated molecules will be synthesized, and tested for activity in inhibiting envelope protein binding to, and dengue virus infection of, target cells. Viruses subvert cell-surface molecules involved in mediating physiological processes to gain access to target cells. In addition to understanding this important element of dengue virus pathogenesis, identification of dengue target cell receptors may also lead to an understanding of the physiological roles of these cellular receptors, as well as advances in understanding interactions between complex carbohydrates and proteins.